The long-term goals of this work are to uncover determinants of inherited and acquired long QT syndrome (LQT), and to make advances that will ultimately allow more drugs to be developed while protecting those at risk from sudden cardiac death. This study focuses on composition and functional analysis of hERG 1a and 1b subunits contributing to channels underlying cardiac IKr, a critical target for inherited and acquired LQT. The specific aims of the current project are to (1) elucidate the developmental and sex-specific profiles of homomeric and heteromeric hERG channels in native tissues, (2) resolve the stoichiometry of these subunits determined by primary sequence in heterologous systems, and that found in native systems, (3) understand the biophysical basis for pharmacological differences previously defined for homomeric and heteromeric channels, and (4) characterize mechanisms of disease specific to the most recently defined hERG 1b isoform. A high-impact component of the proposal is the use of a large sample of normal ventricular tissue from human donors, and the paired analysis of individual specimens with respect to channel composition and physiological response to challenge by drugs that cause acquired long QT syndrome (LQT). Two independent, innovative strategies will be utilized for determining subunit stoichiometry in both heterologous and native tissues. The mechanism underlying differential potency of drugs to homomeric and heteromeric channels will be determined by testing two hypotheses relating to structural and functional differences of the 1a and 1b subunits. How perturbation of the 1b subunit contributes to disease will be evaluated with experiments reporting changes in cellular trafficking or functional alterations caused by novel mutations derived from a proven genomic screening approach, by the evaluation of a genetic modifier persistently associated with LQT populations, and by selective knockdown of the 1b subunit in native tissues. These experiments are expected to elucidate how hERG 1a and 1b subunits contribute to native IKr and how their different expression profiles dictate normal function and susceptibility to long QT syndrome and sudden cardiac death in children, men and women. PUBLIC HEALTH RELEVANCE: If successful, these studies will expand current understanding of the molecular basis for inherited arrhythmias and those caused by block from drugs intended for other therapeutic targets. They will facilitate development of better drug safety screening approaches by providing information of stratified risk for sudden cardiac death among children, women and men.